1. Field of the Invention
The present invention relates to a scanning optical device and an observation method.
This application is based on Japanese Patent Application No. 2007-105568, the content of which is incorporated herein by reference.
2. Description of Related Art
There has been hitherto known a scanning optical device for obtaining a bright fluorescence image having high resolution in the depth direction of a specimen by using a multiphoton excitation phenomenon. Fluorescence emitted from the inside of the specimen is scattered by the specimen, and thus there is a problem that a fluorescence image based on fluorescence generated at a deep position of the specimen is darker than a fluorescence image based on fluorescence generated at a shallow position of the specimen.
In order to solve this problem, for example, according to the invention disclosed in Japanese Unexamined Patent Application, Publication No. 2000-275541, obtained image data are subjected to image processing to correct apparent variation in brightness on a screen.
However, when the correction is made by subjecting obtained image data to the image processing, it induces a problem that the amount of brightness information contained in the image data itself is reduced and thus the corrected fluorescence image is unclear. For example, when a light source, an optical scanning portion, or a photodetector is adjusted so that a fluorescence image based on fluorescence generated at a shallow position of a specimen is clear, a fluorescence image obtained by detecting fluorescence generated at a deep position of the specimen is dark as a whole and has a small amount of brightness information. Therefore, even when the brightness is corrected by the image processing, no clear image can be constructed.
Conversely, when the light source, the optical scanning portion or the photodetector is adjusted so that a fluorescence image based on fluorescence generated at a deep position of the specimen is clear, a fluorescence image obtained by detecting fluorescence generated at a shallow position of the specimen is saturated in brightness, and thus even when the brightness is corrected by the image processing, no clear image can be constructed likewise.
That is, in order to obtain a clear fluorescence image irrespective of the depth of the specimen, it is necessary to obtain a fluorescence image having the same brightness without conducting the image processing.
Furthermore, when an observation position is moved in a horizontal direction with respect to a specimen having an undulating surface, the depth of a focus position in the specimen varies in spite of no variation in the absolute height of the focus position of exciting light at the device side, so that the magnitude of scattering varies.